Application of Pluronic gel to the study of root-knot nematode behaviour

Abstract

Pluronic F-127 is a stable, non-toxic, copolymer that forms a gel at room temperature and a liquid at 15°C when the concentration is 20-30%. Root-knot nematode (Meloidogyne spp.) second-stage juveniles can move freely through the gel and display attraction toward roots of tomato, Medicago truncatula, common bean and Arabidopsis. The excellent clarity of the gel allows examination of behavioural changes in the nematode as it reaches and infects its host. Attraction assays showed that Meloidogyne javanica and M. incognita move to roots much more rapidly than does M. hapla. Nematodes form aggregates when in contact with root tips suggesting that the presence of a signal from the nematode or from the root is involved in the attraction. Nematodes suspended in Pluronic gel without roots aggregate into balls after 1 to 2 days. A coverslip placed on the gel accelerates and serves as a focus for the aggregation, suggesting that lower oxygen, or perhaps a volatile attractant, is involved in this behaviour. These observations demonstrate that Pluronic gel is a useful medium for dissecting attraction of root-knot nematodes to their hosts and for studying additional aspects of their behaviour.

RNAi-induced silencing of an effector confers transcriptional oscillation in another group of effectors in the root-knot nematode, Meloidogyne incognita

“…In order to examine the silencing effect of CWDE and pioneer genes on infectivity of M. incognita, penetration ability of dsRNA-fed J2 in tomato root was analysed in pluronic gel medium, PF-127 (Wang et al, 2009;Dutta et al, 2011). Five ml of 23% PF-127 (Sigma) was pipetted into a 35 mm Petri dish and a 5-day-old tomato (cv.…”

Three dimensional study of wounded plant roots recruiting entomopathogenic nematodes with Pluronic gel as a medium

“…Gel preparation and attraction assays were conducted according to Wang et al (2009b). Approximately 23% (wt/vol) Pluronic F-127 gel (NF Prill Poloxamer 407, BASF, Mt Olive, NJ, USA) in 10 mM Tris-MES (morpholino-ethanesulfonic acid) buffer (Sigma-Aldrich) was made under refrigeration at 4°C and with continuous stirring overnight.…”

“…To optimize the system, nematode Hb-HBN attraction to Chinese chive roots was tested with variable concentrations (200, 400 and 800 IJs/mL) based on the response of root-knot nematodes to plant roots (Wang et al, 2009b). Collected nematodes at different concentrations were washed with sterile water through centrifugation for three times.…”

Effect of chemical stress imposed by Solanum nigrum in calreticulin and beta-1,4- endoglucanase genes and in infectivity of Pratylenchus goodeyi

“…A solution of Pluronic gel (23 %w/v; Pluronic F-127, SIGMA) was prepared by stirring in sterile distilled water to dissolve the powder for 24 h at 4°C (Wang et al 2009). The resulting gel was refrigerated at 4°C until used for nematode behaviour studies.…”

“…A root tip (Rv), 1 cm long, from banana plants cultured in vitro was placed in the middle of a Syracuse (2 cm diameter) containing 1 ml of Pluronic gel. Ten P. goodeyi, mainly adults, were hand-picked and put on the edge of each Syracuse and the Pluronic gel allowed to solidify at room temperature (Wang et al 2009). A Syracuse containing Pluronic gel but without a root was used as control to assess nematode movement.…”